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@ARTICLE{Krmer:256486,
      author       = {Krämer, Christina E. M. and Singh, Abhijeet and Helfrich,
                      Stefan and Grünberger, Alexander and Wiechert, Wolfgang and
                      Nöh, Katharina and Kohlheyer, Dietrich},
      title        = {{N}on-{I}nvasive {M}icrobial {M}etabolic {A}ctivity
                      {S}ensing at {S}ingle {C}ell {L}evel by {P}erfusion of
                      {C}alcein {A}cetoxymethyl {E}ster},
      journal      = {PLoS one},
      volume       = {10},
      number       = {10},
      issn         = {1932-6203},
      address      = {Lawrence, Kan.},
      publisher    = {PLoS},
      reportid     = {FZJ-2015-06382},
      pages        = {e0141768},
      year         = {2015},
      abstract     = {Phase contrast microscopy cannot give sufficient
                      information on bacterial metabolic activity, or if a cell is
                      dead, it has the fate to die or it is in a viable but
                      non-growing state. Thus, a reliable sensing of the metabolic
                      activity helps to distinguish different categories of
                      viability. We present a non-invasive instantaneous sensing
                      method using a fluorogenic substrate for online monitoring
                      of esterase activity and calcein efflux changes in growing
                      wild type bacteria. The fluorescent conversion product of
                      calcein acetoxymethyl ester (CAM) and its efflux indicates
                      the metabolic activity of cells grown under different
                      conditions at real-time. The dynamic conversion of CAM and
                      the active efflux of fluorescent calcein were analyzed by
                      combining microfluidic single cell cultivation technology
                      and fluorescence time lapse microscopy. Thus, an
                      instantaneous and non-invasive sensing method for apparent
                      esterase activity was created without the requirement of
                      genetic modification or harmful procedures. The metabolic
                      activity sensing method consisting of esterase activity and
                      calcein secretion was demonstrated in two applications.
                      Firstly, growing colonies of our model organism
                      Corynebacterium glutamicum were confronted with intermittent
                      nutrient starvation by interrupting the supply of iron and
                      carbon, respectively. Secondly, bacteria were exposed for
                      one hour to fatal concentrations of antibiotics. Bacteria
                      could be distinguished in growing and non-growing cells with
                      metabolic activity as well as non-growing and
                      non-fluorescent cells with no detectable esterase activity.
                      Microfluidic single cell cultivation combined with high
                      temporal resolution time-lapse microscopy facilitated
                      monitoring metabolic activity of stressed cells and
                      analyzing their descendants in the subsequent recovery
                      phase. Results clearly show that the combination of CAM with
                      a sampling free microfluidic approach is a powerful tool to
                      gain insights in the metabolic activity of growing and
                      non-growing bacteria.},
      cin          = {IBG-1},
      ddc          = {500},
      cid          = {I:(DE-Juel1)IBG-1-20101118},
      pnm          = {581 - Biotechnology (POF3-581)},
      pid          = {G:(DE-HGF)POF3-581},
      typ          = {PUB:(DE-HGF)16},
      UT           = {WOS:000363920300081},
      doi          = {10.1371/journal.pone.0141768},
      url          = {https://juser.fz-juelich.de/record/256486},
}